This proposal will investigate the role of oxalate in the pathogenesis of kidney stone disease. A central tenet of our research over the past several years has been that tubular damage/or dysfunction plays a crucial role in the retention of calcium oxalate crystals within the kidney, with damaged cells serving either as sources of membraneous debris for crystal nucleation or binding, and/or as sites for crystal adherence. Our recent studies have demonstrated that oxalate damages renal epithelial by process(es) involving reactive oxygen molecules. Moreover, oxalate toxicity was observed within the range of concentrations that occur normally in urine. Other studies demonstrated effects of oxalate on the proliferation of renal epithelial cells. Thus oxalate may exert dual effects on the viability and growth of renal cells. The present proposal will examine these effects of oxalate and their possible role in kidney stone disease by addressing the following: 1) How does oxalate induce damage to renal tubular cells? Are the effects due to increased production of peroxyl radicals, decreased glutathione, increase lipid peroxidation, and/or alterations in Ca2+i or pHi? 2) Does oxalate stimulate cell proliferation or does the increase in DNA synthesis reflect DNA repair or a prelude to apoptosis? Are these responses also mediated by free radicals? 3) Are the effects of oxalate selective for proximal tubular cells (LLC-PK1 cells) or can we observe similar changes in distal tubular cells (MDCK cells) and/or in inner medullary collecting duct cells (primary cultures)? 4) Can we detect similar oxalate induced changes in viability and growth in renal tubular cells from stone-forming animals? 5) Can we reduce oxalate toxicity by genetic manipulations (transfection with the oxalate oxidase gene to promote oxalate degradation or with the bcl-2 gene to protect against oxidant stress)? Such studies will provide valuable knowledge of specific oxalate-induced changes in renal epithelial cell function and insight as to how such changes might contribute to stone formation. Understanding the cellular basis for oxalate action should provide the basis for new strategies in the prevention and management of urolithiasis.